X-Ray snapshots of a pyridoxal enzyme: a catalytic mechanism involving concerted [1,5]-hydrogen sigmatropy in methionine γ-lyase.

Pyridoxal 5'-phosphate (PLP)-enzymes are essentially involved in amino acid and amine metabolism of a wide variety of organisms. Despite their extensive biochemical studies, there are little evidence and structural data to comprehensively elaborate the catalytic mechanism. We obtained X-ray snapshots of l-methionine γ-lyase from Entamoeba histolytica (EhMGL), a PLP-enzyme catalyzing the γ-elimination reaction of methionine. Here, we suggest a catalytic mechanism of EhMGL by using the X-ray snapshots covering all stages of this multistep catalysis reaction. Initial formation of a Michaelis complex is followed by the migration of double bond from the C4'=Nα-Cα moiety in an intermediate PLP-methionine imine to C4'-Nα=Cα in pyridoxamine 5'-phosphate (PMP)-α,ß-dehydromethionine imine without intervention of a putative quinonoid intermediate. The enzyme can facilitate the subsequent γ-elimination of methanethiol by the possible general acid-base catalysis of Tyr108 for the E1cB mechanism, enabling to form the ene-imine C4'-Nα=Cα-Cß=Cγ structure with the s-cis conformation, which is prerequisite for the non-enzymatic symmetry-allowed suprafacial [1,5]-hydrogen shift to complete the catalytic cycle by releasing α-ketobutyrate. The mechanism based on the X-ray snapshots is consistent with the reactivity of MGL toward methionine analogues. The generality of such a mechanism involving non-enzymatic concerted reaction in other PLP enzymes is discussed.